This research seeks to define the biochemistry and cell biology of endothelial cell basement membrane degradation by malignant cells. Basement membranes elaborated by cultured bovine endothelial cells and labeled with radioactive precursors in specific components will be prepared using novel techniques developed in this laboratory. The subendothelial matrix has been characterized biochemically by sequential enzyme hydrolysis, gel electrophoresis, electron microscopy, and immunofluorescence. The degradation of basement membrane by human tumor cell lines has been investigated in detail and the role of tumor proteases, especially the tumor cell plasminogen activator, defined. Human fibrosarcoma cells are capable of the complete degradation of the subendothelial matrix after removal of the endothelial cells. We also have found that living endothelial cells secrete a factor which inhibits the degradation of the membrane by tumor cells. This regulation of tumor cell aggressive behavior by a normal cell type could have major importance in the metastatic process and we are characterizing it in more detail. Subendothelial matrices also are being used for the routine culture of primary explants of human pediatric tumors. The subendothelial matrix also stimulates the differentiation of normal cells cultured upon it and we are determining which factors within the subendothelium play important roles in this stimulation. The subendothelial matrix, therefore, can be degraded by invasive cells and can modulate the differentiation of certain other cell types, and endothelial cells themselves inhibit protease production by malignant cells. All of these properties may play key roles in the establishment of secondary tumors during metastasis. (A)